RUI: Enzyme Characterization and Gene Cloning of a Putative Drosophila Homolog for Mammalian Dipeptidyl Peptidase IV

University Of San Francisco, San Francisco CA

Investigators

Abstract

Proline specific N-terminal processing is vital to both the activation and/or degradation of many biologically important proteins and peptides. The N- terminal prolyl dipeptidyl peptidase (DPP IV ) has been identified in an impressive array of organisms. These include several species of mammal, from mouse to man, frog, fungi, nematode, flies, sea anemone, and several species of bacteria. DPP IV is involved in the metabolism of peptides during the intestinal digestion and renal transport of polypeptides. It functions in the activation of T-lymphocytes in the immune system of mammals, and the regulation of DNA synthesis, cell proliferation and cytokine production in the hematopoietic system. In some mammals, it functions in signal transduction and cell adherence phenomena. In the human nervous system, it regulates the activity of neuropeptide Y and in the sea anemone is likely to play a role in the processing of at least one neuropeptide (metamorphosin A). In insects, DPP IV is presumed to function in the processing of several peptides, including the immune peptides of several species and the peptide melittin in honeybee venom. However, only a few natural substrates for the enzyme have been confirmed. This laboratory recently reported on a mutant stock (omega) in the fruit fly Drosophila melanogaster that has lost the enzyme DPP IV and as a consequence displays several sublethal defects: A larval developmental delay, a 40% decrease in male fertility and in the ability of males to successfully fertilize females, and a loss of post translational processing of a larval cuticle protein. These defects point to a multiplicity of processes for which the enzyme is required in Drosophila, implying that there are several, as yet unknown, peptides that are processed by DPP IV in addition to the cuticle protein. This research includes plans to clone the DPP IV gene from Drosophila using the available genomic clones of the Berkeley Drosophila Genome Project, one of which has recently been identified as having an homologous sequence imbedded in it. This will allow comparison of its structure with the known DPP IV genes from other organisms. The enzyme will also be purefied. This will allow characterization of its biochemical properties. The role of DPP IV in the processing of Drosophila peptides, in particular with respect to the cuticle proteins, the immune peptides and testicular peptides, will be clarified using available injection and PAGE procedures. These data will lead to a further understanding of the DPP IV function in the innate immune system and in other peptide processing systems of Drosophila, thus further demonstrating the ubiquity of this important enzyme in eukaryotic organisms.